THE ACTION OF A PHORBOL ESTER ON VOLTAGE-DEPENDENT PARAMETERS OF THE SODIUM CURRENT IN ISOLATED HIPPOCAMPAL-NEURONS

The action of a phorbol ester (phorbol 12, 13-diacetate) on the voltage-activated sodium current has been investigated by the voltage-clamp method in acutely isolated pyramidal neurons from rat hippocampus. The intracellular perfusion of isolated pyramidal neurons for 30-40 min induced a gradual 10-15 mV shift in both the current-voltage relationship and voltage-dependent steady-state inactivation to more negative potentials. The application of phorbol ester (1-10 mu M) to isolated neurons for the same time increased the amplitude of sodium current by 15-20%, shifted the above-mentioned voltage-dependent parameters for an additional 10-15 mV in the same direction and changed the slope of the steady-state inactivation curve. In contrast, after prolonged incubation of slices in the phorbol ester-containing solution (1-10 mu M) for 0.5-3 h, subsequent application of phorbol ester at the same concentration caused neither the additional shift of the voltage-dependent characteristics of sodium channels nor the change of the slope of the steady-state inactivation curve. However, in this case an increase in the amplitude of sodium current by 15-20% during 30-40 min intracellular perfusion was observed. Taking into account that long-term incubation of slices in the phorbol ester-containing solution can deplete protein kinase C, we suggest the existence of two types of effects of the phorbol ester on sodium channels: direct action of the phorbol ester, activating protein kinase C (a shift of the voltage-dependent characteristic of sodium channels for 15 mV and the change of the slope of the steady-state inactivation curve), and an indirect action (the increase of the sodium current amplitude by 15-20%). Apparently, the latter effect is mediated by both the binding of phorbol-12,13-diacetate molecules with the cellular membrane and by the intracellular perfusion of the neurons.